Amyotrophic lateral sclerosis (ALS) is a fatal degenerative disorder of motor neurons. It is predominantly sporadic in occurrence, although 10% of cases are inherited as autosomal dominant traits. Mutations in the cytosolic copper-zinc superoxide dismutase gene (SOD1) account for 25% of dominantly-inherited cases. Considerable insight into the pathogenesis of both familial and sporadic ALS has been derived from studies of SOD 1 gene mutations. Several more dominantly-inherited ALS genes have been mapped but not yet identified. The goal of this project is to apply high-throughput genomic methodology systematically to identify novel ALS genes, and to develop new transgenic mouse models of ALS with these genes. These efforts will target the dominant ALS loci that we have recently identified on chromosomes 9, 16, and 20. The Specific Aims of this project are to: (1) Clone the disease loci from affected individuals with chromosome 9-, 16-, and 20- linked ALS in bacterial artificial chromosome (BAC) libraries prepared from mouse-human somatic cell hybrids haploid for the chromosomes of interest; (2) Screen candidate genes from the three ALS loci by exon sequencing; (3) Use high-through-put sequencing to determine the genomic sequence of the BAC contig spanning the ALS loci; (4) Confirm the disease association of candidate mutations; (5) Engineer mouse knock-in models of ALS by microinjection of BACs harboring mutations in the genes identified in Aims 1-4. Significance: These studies will be important because: (A) This project may serve as a new paradigm for the systematic identification of other Mendelian disease traits. (B) These studies will elucidate pathways critical for the survival of motor neurons. The same pathways may be involved in sporadic ALS. (C) An understanding of the molecular pathophysiology of this disease is essential to the rational design of therapeutic interventions. (D) The genes identified in these experiments can be used to create additional animal and cell models of ALS, allowing improved drug design and screening. (E) The characterization of cell death pathways in ALS may provide insights into mechanisms involved in other neurodegenerative disorders such as Alzheimer's and Parkinson's diseases.